3T3-L1 fibroblasts differentiate in culture to resemble adipose cells both morphologically and biochemically. The number of glucose transporters has been measured in subcellular membrane fractions from these cells during differentiation. The data suggest that the glucose transporter undergoes differential processing and that functional, insulin-responsive glucose transporters may be different from the insulin-insensitive (basal) glucose transporter. In a preliminary series of experiments, insulin appears to stimulate glucose transport in isolated human adipose cells by a translocation mechanism similar to that observed in rat adipose cells and diaphragm. Conditions have been established which allow the isolation of rat adipose cell plasma membranes retaining a large part of the stimulatory effect of insulin in intact cells. In these membranes, the magnitude of glucose transport stimulation in response to insulin was compared with the concentration of glucose transporters as measured with the cytochalasin B binding assay or by immunoblotting with an antiserum against the human erythrocyte glucose transporter. The results suggest that in addition to stimulating the translocation of glucose transporters to the plasma membrane, insulin appears to induce a structural or conformational change in the glucose transporter manifested in an altered activation energy for plasma membrane glucose transport and possibly in an altered immunoreactivity as assessed by Western blotting.